Effect of formulation and peptide folding on the fibrillar aggregation, gelation, and oxidation of a therapeutic peptide.

The physical and chemical stability of therapeutic peptides presents challenges in developing robust formulations. The stability of the formulation affects product safety, efficacy and quality. Therefore, an understanding of the effects of formulation variables on the peptide's conformational structure and on its possible physical and chemical degradation is vital. To this end, computational and experimental analysis were employed to investigate the impact of formulation, peptide folding and product handling on oxidation, fibrillar aggregation and gelation of teriparatide. Teriparatide was used as a model drug due to the correlation of its conformation in solution with its pharmacological activity. Fibrillar aggregation and gelation were monitored using four orthogonal techniques. An innovative, automated platform coupled with ion mobility mass spectrometry was used for profiling chemical degradants. Increases in teriparatide concentration, pH, and ionic strength were found to increase the rate of fibrillar aggregation and gelation. Conversely, an increase in peptide folding and stabilization of the folded structures was found to decrease the rate of fibrillar aggregation and gelation. Moreover, the rate of oxidation was found to be inversely related to its solution concentration and extent of peptide folding. The present study provides an insight into formulation strategies designed to reduce the potential risk of physical and chemical degradation of peptides with a defined conformation.

Zinc supplementation improves the harvest purity of ß-glucuronidase from CHO cell culture by suppressing apoptosis.

The variability of trace metals in cell culture media is a potential manufacturing concern because it may significantly affect the production and quality of therapeutic proteins. Variability in trace metals in CHO cell culture has been shown to impact critical production metrics such as cell growth, viability, nutrient consumption, and production of recombinant proteins. To better understand the influence of excess supplementation, zinc and copper were initially supplemented with 50-µM concentrations to determine the impact on the production and quality of ß-glucuronidase, a lysosomal enzyme, in a parallel bioreactor system. Ethylenediaminetetraacetic acid (EDTA), a metal chelator, was included as another treatment to induce a depletion of trace metal bioavailability to examine deficiency. Samples were drawn daily to monitor cell growth and viability, nutrient levels, ß-glucuronidase activity, and trace zinc flux. Cell cycle analysis revealed the inhibition of sub-G0/G1 species in zinc supplemented cultures, maintaining higher viability compared to the control, EDTA-, and copper-supplemented cultures. Enzyme activity analysis in the harvests revealed higher specific activity of ß-glucuronidase in reactors supplemented with zinc. A confirmation run was conducted with supplementations of zinc at concentrations of 50, 100, and 150 µM. Further cell cycle analysis and caspase-3 analysis demonstrated the role of zinc as an apoptosis suppressor responsible for the enhanced harvest purity of ß-glucuronidase from zinc-supplemented bioreactors.

Quality attributes and evaluation of pharmaceutical glass containers for parenterals.

Pharmaceutical containers for parenterals have been predominantly manufactured using glass as a packaging material of choice, especially Type-I glass, since it has been regarded as a chemically inert and an effective container closure system (CCS). Nevertheless, there have been reports and recalls related to glass quality issues, such as breakage, flakes, and particles observed in marketed products. The novelty of this research is based on the knowledge gathered from our previously conducted risk assessments and establishing a comprehensive testing platform focused on risk factors for glass container failure modes and applicability to other types of pharmaceutical containers. The evaluation of container quality attributes was performed for three model glass vials using a mechanical and chemical durability testing platform: freeze-thaw, lyophilization, compression, scratch tests; visual inspection, pH, particle size analyses, extractable, leachable and imaging studies that were conducted under normal (4 and 25 °C), and stress condition (60 °C), respectively. The performance between the glass containers tested under certain stress conditions (failure modes) were variable and differentiated. The systematic platform testing approach shows the importance of lab-based risk evaluation in assessing common failure modes of pharmaceutical containers, since the quality attributes for injectable products are complex and can impact final product quality.